Control LNCaP cells (shScr) and NPM1 knocked-down LNCaP cells (shNPM1) were seeded in 24-wells plates and grown to confluence for 24 hours. The monolayer culture was then scrape-wounded with a sterile micropipette tip in order to create a gap of constant width. Cellular debris were washed with Phosphate Buffered Saline 1× (PBS) (Life Technologies). Cells were next grown in RPMI 1640 10%FBS that was replaced 12 hours after wounding and then every 24 hours. LNCaP cell migration was photographed into the wounded region at 24, 48 and 72 hours following the scraping (100× magnification) and remaining wound areas were then quantified with ImageJ free software.
Phosphate buffered saline 1 pbs
Manufactured by Thermo Fisher Scientific
Sourced in United States
Phosphate buffered saline (1× PBS) is a widely used aqueous solution that maintains a stable pH and ionic strength. It is commonly used as a buffer in various laboratory applications, including cell culture, immunoassays, and biochemical experiments.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Nageotte counting chamber, by Hausser Scientific (1 mentions)
Celltracker green, by Thermo Fisher Scientific (1 mentions)
Hemocytometer, by Hausser Scientific (1 mentions)
Polyvinyl alcohol (pva), by Fujifilm (1 mentions)
Hydrochloric acid (hcl), by Fujifilm (1 mentions)
Aniline, by Fujifilm (1 mentions)
Ammonium persulfate, by Fujifilm (1 mentions)
Cacl2 2h2o, by Merck Group (1 mentions)
Texas red conjugated dextran, by Thermo Fisher Scientific (1 mentions)
Sodium chloride (nacl), by Merck Group (1 mentions)
Potassium chloride (kcl), by Merck Group (1 mentions)
Sucrose, by Merck Group (1 mentions)
Hepes, by Merck Group (1 mentions)
Nahco3, by Merck Group (1 mentions)
Mgcl2, by Merck Group (1 mentions)
Trehalose, by Merck Group (1 mentions)
Lsm 900, by Zeiss (1 mentions)
Methyl alcohol hplc grade, by Merck Group (1 mentions)
9 protocols using phosphate buffered saline 1 pbs
1
Quantifying LNCaP Cell Migration
2
Cancer Cell Spiking Protocol
3
Ammonium Persulfate Functionalization Protocol
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Ammonium persulfate (APS), hydrochloric acid (HCl), aminophenyl boronic acid (APBA), aniline (ANI), polyvinyl alcohol (PVA), and glucose were purchased from Wako Pure Chemical Industries, Ltd. Phosphate-buffered saline (1× PBS, pH 7.4) was purchased from Thermo Fisher Scientific Inc.
4
Dextran Uptake Assay for Nephrocyte Function
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The dextran uptake assay was used to measure nephrocyte filtration function ex vivo. Nephrocytes were dissected from 4-day-old adult flies (females) and kept in artificial hemolymph [70 mmol/l NaCl (Carolina), 5 mmol/l KCl (Sigma), 1.5 mmol/l CaCl2·2H2O (Sigma-Alrich), 4 mmol/l MgCl2 (Sigma-Aldrich), 10 mmol/l NaHCO3 (Sigma-Aldrich), 5 mmol/l trehalose (Sigma), 115 mmol/l sucrose (Sigma-Aldrich), and 5 mmol/l HEPES (Sigma-Aldrich), in water]. Cells were incubated with Texas Red-conjugated dextran, 10,000 MW (0.05 mg/ml; Invitrogen) for 20 min, and then fixed with 4% paraformaldehyde in phosphate-buffered saline (1× PBS) (Thermo Fisher Scientific) for 10 min. Dextran uptake capacity was based on nephrocyte fluorescence levels, assayed by fluorescence confocal microscopy (ZEISS LSM 900; see details below). For quantification, 30 nephrocytes from six female adult flies were analyzed per genotype. The results are presented as mean±s.d.
5
Isolation of Intestinal Epithelial Exosomes
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Intestinal epithelial cells were transfected with poly I:C (0.1, 1, 10 µg/ml) for 4 h and fresh-culturing medium containing 10% exosome-free FBS was added. At 48 h post-transfection, IECs supernatant (SN) was collected and exosomes were isolated through multiple rounds of centrifugation and filtration as previously reported (24 (link)). Briefly, 10 ml of SN were centrifuged at 300 × g for 10 min to remove floating cells, then at 2,000 × g for 10 min, and 10,000 × g for 30 min to remove cell debris, shedding vesicles, and apoptotic bodies. Finally, exosomes pellet were collected by ultracentrifugation at 100,000 × g for 70 min. For further purification, the pellets were washed with phosphate buffered saline (1× PBS) (Gibco, NY, USA) and centrifuged at 100,000 × g for 70 min. The pellet was resuspended in 100 μl 1× PBS, then immediately stored at −80°C until use.
6
Heteroclitic Peptides with Novel p4 Substitutions
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Heteroclitic peptides (htcPep) with substitution in the p4 residue were described in our previously study (17 (link)). An additional htcPep was designed introducing at p4 a non-natural aa residue based on the Tryptophan structure (NAL). Alternatively, a peptide library including 20 peptides was prepared by random introduction of all 20 possible amino acids at position 4 (MIX). Individual htcPep were synthesized at a purity > 95% determined by LC-MS analyses. Lyophilized powder was dissolved in dimethylsulfoxide (DMSO; Sigma-Aldrich), diluted in phosphate-buffered saline (1× PBS; Gibco Life Technologies) and stored at − 80°C until use.
7
CalliSpheres Evaluation for Drug Delivery
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CalliSpheres® (Callisyn Biomedical-Suzhou, Inc., Suzhou, China) with sizes of 50–150, 100–300, and 300–500 μm were evaluated. Each package contains 1 g of DEB and 7 of mL normal saline, making up a total volume of about 8 mL. Idarubicin hydrochloride for injection (National Medicine Permission Number H20203345, Batch No. 200917) and iopamidol injection (National Medicine Permission Number H20203293, Batch No. 200812) were kindly provided by Chia Tai Tianqing Pharmaceutical Group Co. Ltd., Nanjing, China. Each IDA package contains hydrochloride powder with 10 mg of active pharmaceutical ingredients (APIs). Idarubicin hydrochloride standard (>99%) was purchased from the National Institutes for Food and Drug Control, China. Methyl alcohol (HPLC grade) was purchased from Sigma-Aldrich, Saint Louis, MO, USA. Phosphate buffered saline (1 × PBS, pH = 7.2~7.4) and fetal bovine serum (FBS) were purchased from Gibco, Grand Island, NY, USA. The 0.9% sodium chloride for injection (0.9% NaCl) and the 5% glucose for injection (5% Glu) were purchased from Shijiazhuang No. 4 Pharmaceutical, China. Ultrapure water (H2O) was obtained from a Millipore system (Millipore, Bedford, MA, USA). All other chemicals were of analytical grade.
8
Anticancer Compound Screening Protocol
9
Characterization of Metal-Metformin Complexes
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The following reagents and solvents were commercially available and used as supplied without further purification: metformin hydrochloride U. S. P. (Metf) (Harman Finochem Limited); CoCl2·6H2O (99%, Sigma-Aldrich); CuCl2·2H2O (99%, Sigma-Aldrich); NiSO4·6H2O (99%, Sigma-Aldrich); and ZnCl2·6H2O (99%, Sigma-Aldrich). Chemical and reagents used in cell culture included phosphate-buffered saline 1× (PBS) (Gibco, Carlsbad, CA, USA), low glucose Dulbecco's Modified Eagle's Medium (LGDMEM) (Sigma-Aldrich, St. Louis, MO, USA), high glucose Dulbecco's Modified Eagle's Medium (HGDMEM) (Sigma-Aldrich, St. Louis, MO, USA), RPMI 1640 Medium (Sigma-Aldrich, St. Louis, MO, USA), fetal bovine serum (FBS) (Invitrogen, Carlsbad, CA, USA), penicillin–streptomycin (P/S) 10 000 U mL−1 (Invitrogen, Carlsbad, CA, USA), glutamine (Gibco, Carlsbad, CA, USA), and the methyl thiazole tetrazolium (MTT) assay (Amresco, Solon, OH, USA).
Thermogravimetric analysis (TGA) was performed in a TA instrument (Discovery SDT 650) under the following conditions: 25–900 °C temperature range, under nitrogen (100 mL min−1 flow) atmosphere, and 20 °C min−1 heating rate. Fourier-transform infrared spectrophotometry (FTIR) spectra were recorded from KBr pellets in the 4000–250 cm−1 range on a Shimadzu IRAffinity-1.
Thermogravimetric analysis (TGA) was performed in a TA instrument (Discovery SDT 650) under the following conditions: 25–900 °C temperature range, under nitrogen (100 mL min−1 flow) atmosphere, and 20 °C min−1 heating rate. Fourier-transform infrared spectrophotometry (FTIR) spectra were recorded from KBr pellets in the 4000–250 cm−1 range on a Shimadzu IRAffinity-1.
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